Department of Pharmaceutical Analysis, College of Pharmacy, Chung-Ang University, Dongjak-Gu, Seoul 156-756, South Korea.
J Chromatogr A. 2010 Jun 25;1217(26):4347-54. doi: 10.1016/j.chroma.2010.04.047. Epub 2010 Apr 22.
Saikosaponins are triterpene saponins derived from the roots of Bupleurum falcatum L. (Umbelliferae), which has been traditionally used to treat fever, inflammation, liver diseases, and nephritis. It is difficult to analyze saikosaponins using HPLC-UV due to the lack of chromophores. Therefore, evaporative light scattering detection (ELSD) is used as a valuable alternative to UV detection. More recently, a charged aerosol detection (CAD) method has been developed to improve the sensitivity and reproducibility of ELSD. In this study, we compared CAD and ELSD methods in the simultaneous analysis of 10 saikosaponins, including saikosaponins-A, -B(1), -B(2), -B(3), -B(4), -C, -D, -G, -H and -I. A mixture of the 10 saikosaponins was injected into the Ascentis Express C18 column (100 mm x 4.6 mm, 2.7 microm) with gradient elution and detection with CAD and ELSD by splitting. We examined various factors that could affect the sensitivity of the detectors including various concentrations of additives, pH and flow rate of the mobile phase, purity of nitrogen gas and the CAD range. The sensitivity was determined based on the signal-to-noise ratio. The best sensitivity for CAD was achieved with 0.1 mM ammonium acetate at pH 4.0 in the mobile phase with a flow rate of 1.0 mL/min, and the CAD range at 100 pA, whereas that for ELSD was achieved with 0.01% acetic acid in the mobile phase with a flow rate at 0.8 mL/min. The purity of the nitrogen gas had only minor effects on the sensitivities of both detectors. Finally, the sensitivity for CAD was two to six times better than that of ELSD. Taken together, these results suggest that CAD provides a more sensitive analysis of the 10 saikosaponins than does ELSD.
柴胡皂苷是从柴胡（伞形科）的根部提取的三萜皂苷，传统上用于治疗发热、炎症、肝病和肾炎。由于缺乏生色团，使用高效液相色谱-紫外（HPLC-UV）分析柴胡皂苷非常困难。因此，蒸发光散射检测（ELSD）被用作紫外检测的有价值替代品。最近，发展了一种带电气溶胶检测（CAD）方法，以提高 ELSD 的灵敏度和重现性。在这项研究中，我们比较了 CAD 和 ELSD 方法在同时分析 10 种柴胡皂苷中的应用，包括柴胡皂苷-A、-B(1)、-B(2)、-B(3)、-B(4)、-C、-D、-G、-H 和 -I。将 10 种柴胡皂苷的混合物注入 Ascentis Express C18 柱（100mmx4.6mm，2.7μm）中，进行梯度洗脱，并通过分流进行 CAD 和 ELSD 检测。我们研究了各种可能影响检测器灵敏度的因素，包括添加剂的各种浓度、流动相的 pH 值和流速、氮气的纯度以及 CAD 范围。灵敏度是基于信噪比来确定的。对于 CAD，在流动相中添加 0.1mM 乙酸铵（pH4.0），流速为 1.0mL/min，CAD 范围为 100pA 时，灵敏度最佳；对于 ELSD，在流动相中添加 0.01% 乙酸，流速为 0.8mL/min 时，灵敏度最佳。氮气的纯度对两种检测器的灵敏度只有很小的影响。最后，CAD 的灵敏度比 ELSD 高 2 到 6 倍。综上所述，CAD 为 10 种柴胡皂苷提供了比 ELSD 更灵敏的分析。
